Abstract The worldwide incidence of breast cancer is 1.68 million. Triple-negative breast cancers (TNBC) account for ~20% of total breast cancer cases (336,000 worldwide). Annually, over 40,000 new cases of TNBC are diag- nosed in the US alone. The treatment options for TNBC patients are limited. Triple negative breast cancer (TNBC) includes breast cancers that lack estrogen receptor, progesterone receptor, and HER-2 proteins. Lack of known targets and patient-to-patient variation of target antigens make TNBC one of the most challenging cancers for developing an effective therapy. New therapies are critically needed, as TNBC is also resistant to checkpoint blockade inhibition alone. We propose to develop an immunotherapy administered in combination with checkpoint blockade to treat metastatic TNBC using an approach that is personalized, thus incorporating patient-specific neoantigens and other tumor antigens that are upregulated or altered in glycosylation pattern, such as MUC-1. Metaclipse?s immunotherapy (autologous therapeutic vaccine) consists of Tumor Membrane Vesicles (TMVs) made from patient-specific tumor tissue, which carry membrane associated tumor antigens and also antigenic epitopes derived from cytosolic proteins as MHC associated peptides. These TMVs are then directly conjugated to potent glycolipid-anchored immunostimulatory molecules (GPI-ISMs) by a novel protein transfer technology. The novelty of the proposed vaccine lies not in the use of ISMs, but in the way they are employed. This direct physical linkage of TMVs and GPI-ISMs allows for simultaneous delivery of the patient?s unique tumor antigen signature and ISMs at the vaccination site to induce a robust antitumor immune re- sponse. Since the TMV vaccine is prepared from whole tumor tissue encompassing not only patient specific variation but also all of the heterogeneity of the tumors. A key advantage of our approach is that the immuno- therapy product can be prepared within a week, which is critical during treatment of aggressive cancers such as TNBC. The company has successfully completed proof-of-concept studies using an immune checkpoint in- hibitor (ICI) resistant pre-clinical mouse model of TNBC. The results demonstrate that immunization with TMV vaccine sensitizes the ICI resistant TNBC to become responsive to ICI therapy. The SBIR direct Phase II award is requested to support further IND enabling preclinical studies, which will facilitate IND meetings with the FDA to advance the TMV vaccine product to the clinic. The following are specific aims of the SBIR direct Phase II grant: (1) determine optimal dose and dosing schedule of the immunotherapy product in inducing an effective antitumor immune response, alone and in combination with immune checkpoint blockade therapy; (2) evaluate safety of the administered immunotherapy product in mice; and (3) prepare for GMP production of human GPI-ISMs and establish benchmarks for a GMP-compliant TMV vaccine product. The proposed plan represents Metaclipse?s critical path for product development to enter into a Phase 1 clinical trial.